Investigation and characterization of viruses and phytoplasmas infecting fig trees

Early detection of fig mosaic disease (FMD) is considered as a fundamental step for an appropriate disease management through the production of pathogen-free plant material. Accordingly, singleplex and multiplex-TaqMan RT-PCR assays were developed to detect single and multiple infections of seven fig viruses, i.e. fig leaf mottle-associated virus 1 (FLMaV-1), fig leaf mottle-associated virus 2 (FLMaV-2), fig mild mottle-associated virus (FMMaV), fig mosaic virus (FMV), fig latent virus 1 (FLV-1), fig cryptic virus 1 (FCV-1) and fig fleck-associated virus (FFkaV). The newly designed primers and probes were constructed upon alignment of viral nucleotide sequences of different isolates retrieved from GenBank. The new techniques were validated and found to be more sensitive and reliable than RT-PCR. Multiplex PCR was able to detect with high efficiency up to five fig viruses simultaneously. These two diagnostic techniques can be useful to control and limit fig virus spread and to support quarantine and certification programs. On the other hand, besides to many viral and fungal diseases, nothing was reported before on phytoplasma infections in fig plants. A small-scale survey was therefore conducted in a fig plot for varietal collection (Locorotondo, South of Italy) to investigate the possible presence of phytoplasmas. Forty-three samples were collected and subjected to PCR and nested PCR assays using universal primers that amplify the 16S rDNA gene. Twenty plants were shown positive to phytoplasma. Sequence analysis generated three sequences sharing the highest similarity with ‘Ca. P. asteris’ and ‘Ca. P. solani’, belonging to the 16SrI and 16SrXII ribosomal groups, respectively. The presence of two phylogenetically distinct phytoplasmas was further verified with group specific qPCR assays and RFLP with TruI conducted on 16S rDNA amplified with M1/M2 and fU5/rU3 primers. This is the first record of phytoplasma occurrence in fig plants.